The ability of viruses to successfully infect the host cell relies to a large degree upon ordered expression of viral genes. Due to their compact genome size, many viruses that rely upon the host RNA polymerase II for transcription contain complex transcription units in which multiple mRNAs are derived from a single primary transcript. The goals of this project are to determine the mechanism by which two families of viruses, adenoviruses and retroviruses, regulate gene expression post- transcriptionally in such complex transcription units. The adenovirus major late transcription unit encodes five families of mRNAs, L1 through L5, whose expression is regulated through poly(A) site selection such that early in the infection use of the L1 site predominates, whereas late all the sites are used roughly equivalently. It has been shown that sequences upstream and downstream of the core 3' end processing signals of the L1 poly(A) site are involved in this regulation. In the proposed studies, the mechanism of action of these sequences during infection will be examined. In vitro studies, using transcription/processing and processing reactions, will be used to identify the factors that respond to these regulatory signals. In vivo systems will be used to investigate the role of adenovirus DNA replication in regulation, as it has been shown that regulation late in infection requires transcription from a replicated template. In retroviruses, sequences regulating poly(A) site choice have also been identified upstream of core processing signals. The role of these sequences will be determined using the same types of in vivo and in vitro systems. It has also been reported that proximity of a poly(A) site to the promoter in the 5' long terminal repeat results in a decreased level of steady state RNA processed at that site. The exact effect of promoter proximity will be studied by examination of nuclear events in retroviral RNA metabolism, taking advantage of recombinant adenoviruses. Together, these studies will increase our understanding of how these viruses use post-transcriptional mechanisms to control gene expression during replication.